Antibiotic Resistance In Bacteria

How Researchers Are Fighting Back

by Laura Schocker

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The 1928 discovery of penicillin has been widely touted as one of the greatest scientific achievements in the past century. This first antibiotic went into more frequent use after World War II, but bacteria quickly found a natural way to ward it off; within two to three years of its introduction into health clinics, scientists isolated the first penicillin-resistant bacteria. Today, pathogens like MRSA (methicillin-resistant Staphylococcus aureus) cause thousands of deaths each year in this country. With experts recognizing that the overuse of all antibiotics is creating a major public health problem, researchers in the lab of Erik Sontheimer have taken a step toward outmaneuvering resistant bacteria.

Antibiotic resistance occurs naturally and is amplified through the process of natural selection, says Sontheimer, associate professor of biochemistry, molecular biology, and cell biology. Using, and especially overusing, antibiotics will kill off bacteria without resistance, promoting the selection and growth of bacteria that do carry resistance. At first, doctors simply prescribed different antibiotics, but in the long run, this, too, can create serious problems. “As more and more selective pressure is applied, you end up getting these super bugs that are resistant to multiple antibiotics,” Sontheimer says.

Sontheimer and post-doctoral fellow Luciano Marraffini became involved in the problem by trying to answer questions about horizontal gene transfer, which is how bacteria can spread antibiotic resistance. In conjugation—one form of horizontal gene transfer—bacteria mate and pass on resistance genes from one cell to the other. This can happen between bacteria that are harmless to humans and those that can be dangerous. Sometimes foreign genes benefit the recipient bacterium, but in other cases (such as when the foreign genes come from viruses) they do not. Therefore many bacteria have evolved mechanisms to limit or control their exposure to horizontal gene transfer. Marraffini and Sontheimer’s findings, which were published in the December 2008 issue of Science, offer insights into a new mechanism used by bacteria to stop horizontal gene transfer.

Understanding Sontheimer and Marraffini’s work requires a quick trip back to high school biology. DNA is made up of two strands of nucleic acids, which are strings of bases labeled with an A, T, C or G. A and T always pair with each other, as do G and C, Sontheimer explains by drawing out the possible combinations on a whiteboard. If you have a nucleic acid with a particular sequence of A, T, C and G, the complementary nucleic acid will pair with it. Over the past decade, scientists have discovered a pathway called RNA interference, which uses this process to control whether or not particular genes are active, including those from viruses. The real-life implications are that drug companies can now focus on finding strands of interfering RNA that are complementary to major viruses, such as HIV or hepatitis, effectively blocking viral replication.

While his lab has been working with RNA interference for the past seven years, the problem is that it only works with organisms that have cells with nuclei, including humans, plants and animals—but not bacteria. In 2007, a major food company published a paper identifying regions within the bacterial genome called CRISPR sequences that function in a way that is conceptually similar to RNA interference to protect bacterial cells from virus infection. The analogies to RNA interference made this topic a natural fit for Sontheimer’s lab.

Marraffini earned his PhD from the University of Chicago in 2008 and then came to Northwestern to work on understanding how CRISPR sequences can stop horizontal gene transfer between bacteria, specifically in terms of antibiotic resistance. Many of the genes that transfer antibiotic resistance are found in plasmids, which are extra bits of DNA separate from the bacterial chromosome. These plasmids can be transferred through conjugation. Sontheimer and Marraffini ultimately concluded that CRISPR sequences can effectively block the transfer of plasmids that carry antibiotic resistance, apparently by targeting the plasmid DNA itself. Without the CRISPR, two bacteria will mate and a copy of the plasmid will be transferred from one to the other. If the CRISPR is present in the recipient cell, there is a sort of bacterial “safe sex,” where the two mate but don’t transfer the plasmid. The basic science in Sontheimer’s lab is focused on the fundamental step of understanding exactly how CRISPR has this effect. “First we have to know very well how it works naturally,” Marraffini says. “Then we can try to engineer it as we want.”

Eventually, researchers might be able to exploit the understanding of CRISPR to prevent the spread of antibiotic resistance, or maybe even destroy resistance that already exists by engineering CRISPR sequences to evict antibiotic resistance plasmids that are already present. “It wouldn’t be the same as developing a new antibiotic,” Sontheimer says, “but the vision is that you might be able to make that cell sensitive to an old antibiotic.” And the findings don’t just apply to antibiotic resistance—horizontal gene transfer can also spread virulence factors, which turn a harmless strain of bacteria into a pathogenic one.

In the meantime, Marraffini and Sontheimer are working on Northwestern’s north campus with “lots of Petri dishes,” experimenting with bacteria and manipulating CRISPR sequences. They can generate useful strains of bacteria by applying recombinant DNA technology and then leave them to mate with each other overnight. The next day, they check whether or not the plasmids transferred by looking for colonies on the dishes.

If their work eventually leads to successful clinical applications, patients in hospitals may someday be able avoid infections like MRSA altogether. It could also lead to fighting antibiotic resistance in other harmful bacteria, including those that cause salmonella, tuberculosis, and cholera. “If this could be exploited like RNA interference, it would be a very, very big thing,” Sontheimer says.

Laura Schocker recently received her master’s degree in journalism from Medill. She specializes in science writing.